Growth of the placenta is essential for a successful human pregnancy. The surface of the human placenta is formed by the syncytiotrophoblast, which is a large multinucleate highly differentiated cell that is responsible for transport of nutrients from the maternal to fetal circulations. Being syncytial in nature, the syncytiotrophoblast does not proliferate and does not independently increase cell size. The growth of the placental surface is dependent on continuous intercellular fusion between the underlying proliferative villous cytotrophoblast layer and the syncytiotrophoblast. Some forms of pregnancy loss and intrauterine growth restriction (IUGR) result from interference with intertrophoblast fusion. However, the mechanism underlying most cases of recurrent pregnancy loss and IUGR is unknown. It would be reasonable to predict that defective intercellular fusion may be the underlying cause of other forms of pregnancy loss or IUGR. It is difficult to investigate this hypothesis because the specific process by which trophoblast fuse is not well understood. From the work of many investigators it appears that intertrophoblast fusion is dependent on three changes in the plasma membrane: the externalization of multiple intercellular adhesion proteins, the expression of one or more fusion proteins, and efflux of aminophospholipids. A few specific changes in fusion-related plasma membrane proteins have been investigated, although this has been done in a piece-meal fashion. There is no information on the total change in relevant plasma membrane proteins related to intercellular fusion. This R21 application is designed to obtain global information about plasma membrane changes related to syncytiotrophoblast fusion using subcellular proteomics. The data obtained from studying changes in the plasma membrane proteome will be confirmed using whole cell proteomics, immunohistology, Western blot analysis, and other techniques. The transcription of fusion- related plasma membrane proteins will be investigated using quantitative RT-PCR. Later studies beyond this proposal will ascertain the importance of individual proteins to the intercellular fusion process. These studies will, for the first time, identify a family of plasma membrane proteins that are essential for intertrophoblast fusion and form the basis for future studies on alterations of the expression of these proteins related to recurrent pregnancy loss and IUGR. PUBLIC HEALTH RELEVANCE: Although intercellular fusion between cytotrophoblast cells is essential for normal growth and function of the human placenta, the normal process and aberrations in that process leading to pregnancy loss or intrauterine growth retardation are understood very little. This proposal uses state-of-the-art techniques, such as subcellular proteomics, to characterize the normal intercellular fusion process and provide a foundation for extensive investigations of pregnancy complications that may arise from defective fusion.